Pregnancy is a unique event in which a fetus develops in the uterus despite being genetically and immunologically different from the mother, and the underlying mechanisms remain poorly understood. To analyze the differential gene expression profiles in nonpregnant and 7 days post coitus (dpc) pregnant uterus of mice, we performed a global proteomic study by 2‐D gel electrophoresis (2‐DE) and MALDI‐TOF‐MS. The uterine proteins were separated using 2‐DE. Approximately 1,000 spots were detected on staining with Coomassie brilliant blue. An image analysis using Melanie III (Swiss Institute for Bioinformatics) was performed to detect variations in protein spots between pregnant and nonpregnant uterus. Twenty‐one spots were identified as differentially expressed proteins, of which 10 were up‐regulated proteins such as alpha‐fetoprotein, chloride intracellular channel 1, transgelin, heat‐shock protein beta‐1, and carbonic anhydrase II, while 11 were down‐regulated proteins such as X‐box binding protein, glutathione S‐transferase omega 1, olfactory receptor Olfr204, and metalloproteinase‐disintegrin domain containing protein TECADAM. Most of the identified proteins appeared to be related with catabolism, cell growth, metabolism, regulation, cell protection, protein repair, or protection. Our results uncovered key proteins of mouse uterus involved in pregnancy.

본 연구는 생쥐 자궁에 있어서 아라키돈산으로부터 prostaglandin의 생성에 관여하는 것으로 추측되는 acyl-CoA sytnhetase 4 유전자의 임신단계별 발현을 확인하고자 실시하였다. Acyl-CoA sytnhetase 4 유전자는 착상 전에는 발현이 증가하는 경향을 나타내었으며 착상 후에는 감소하였다. 이러한 발현의 양상은 세포막의 인질로부터 아라키돈산을 유리시키는 cPLA2의 발현과 유리된 아라키돈산으로부터 prostaglandin의 생성에 관여하는 COX1과 COX2의 발현 양상과 일치하였다. 이러한 결과는 세포막에서 유리된 아라키돈산이 무한적으로 COX1과 COX2에 의하여 prostaglandin의 생성에 이용되는 것이 아니라 acyl-CoA sytnhetase 4에 의하여 세포막의 인지질로 되돌려져 prostaglandin의 생성을 조절하는 기능을 세포가 수행하고 있는 것으로 추정된다.

CREBZF (cAMP-response element binding protein zhangfei) is a member of ATF/CREB family, and which regulates various cellular functions by suppressing major factors with direct interaction. In this study, we have examined the expression of CREBZF on mouse endometrium during uterus estrous cycles and estrogen (E2) treatment. In uterus, CREBZF mRNA expression was higher than other organs and mRNA and protein of CREBZF was increased in proestrus phase and decreased in estrus phase. The expression of CREBZF in 3-weeks old mouse uterus was reduced by E2 injection in endometrium. In addition, the expression of progesterone receptor, a marker of E2 in ovariectomized mice was found to be strongly expressed in stroma, while CREBZF was only expressed in epithelium. Also, we conformed that E2-suppressed CREBZF was restored by co-injection of ICI 182,780, an estrogen receptor antagonist. Overall, these results suggest that CREBZF is regulated by estrogen and involved in ER signaling pathway in mouse uterus.

The Hippo signaling pathway is essential for regulating proliferation, differentiation, and apoptosis in mammalian cells. Hippo signaling pathway exists in most body tissues and organs, where it controls the size of organs and tissues by keeping cell growth in check and promoting cell death as needed. It has been reported that the members of Hippo signaling pathway are highly expressed in mammalian ovaries and uteri. However, the regulatory mechanism of this pathway in the uterus during estrous cycle regulation remains unclear. Serine/Threonine Protein Kinase 4 (STK4, also known as MST1, a homolog of Hippo in Drosophila) is a major factor of Hippo signaling pathway. However STK4 in the mouse uterus has not yet been examined. The purpose of our study was to determine the expression of STK4 during the estrous cycle and regulation by estrogen in the mouse uterus. We found that STK4 was dynamically expressed in uterine endometrium during the estrous cycle. STK4 highly expressed at the estrus, diestrus, and were found to dramatically decrease as it progressed to the proestrus, metestrus stage of uterus during the estrous cycle. Expression of STK4 was dominant in glandular epithelial and luminal epithelial of proestrus, estrus, and diestrus stage, whereas in metestrus stage, expression of gene intensity was faint. Estrogen or estrogen receptor antagonist ICI 182,780 treatment, in ovariectomized mouse uterus, Expression of STK4 and its downstream genes were increased by estrogen. Our results show that the Hippo signaling pathway is estrogen-dependent in the mouse uterus. These informations will give us on sights to understand uterine dynamics during the estrous cycle.

Autophagy is a major cellular catabolic pathway and is tightly associated with survival and death of cells. The involvement of autophagy during prolonged survival of blastocysts in the uterus is established and it was assumed that ovarian steroid hormones – estrogen (E2) and progesterone (P4) – play important roles in its regulation. The uterus is a major target organ of E2 and P4. To examine if E2 or P4 modulate autophagy in the mouse uterus in vivo, the following three systems were used. 1) Normal pregnancy model (days 1 to 8); 2) delayed implantation model; 3) ovariectomized (OVX) mice model treated with single steroid hormone. Six-week-old virgin ICR mice were used for pregnancy and OXV. OVX mice received P4 (1 mg/0.1 ml) or E2 (100 ng/0.1 ml) after 12 days of rest. Collected uteri were subjected to Western blotting and immunofluorescence staining using anti-LC3B antibody to monitor autophagy. In pregnant mouse uterus, the autophagic response was downregulated after implantation. In OVX model, either E2 or P4 injection downregulated the autophagic response in the uterus within several hours. To confirm whether hormone-induced downregulation is mediated by classical estrogen receptor (ER) and progesterone receptor (PR), receptor antagonists (ICI 182,780 and RU-486) were co-treated. Antagonist-treated uteri showed recovery of autophagic response, suggesting that ER or PR mediates hormonal effects on autophagy. In oder to determine which signaling pathway is involved in autophagic regulation by E2, rapamycin (5 mg/kg), a mTOR inhibitor, and LY294002 (5 mg/kg), a PI3 kinase inhibitor, were used. Rapamycin and LY294002 were injected just before E2 injection to OVX mice. Western blotting was performed by using anti-phospho-mTOR and anti-AKT antibodies. We observed that rapamycin treatment partially antagonized downregulation of autophagic activation by E2, whereas LY294002 treatment did not have any effect. Therefore, downregulation of autophagy by E2 seems to be partially mediated by mTOR pathway. Collectively, this study suggests that ovarian steroid hormones are upstream controllers of autophagic response in the mouse uterus.

시상하부에서 생성되는 nesfatin-1/NUCB2가 음식물 섭취와 에너지 대사를 조절한다는 것이 보고된 이후로, 최근 생쥐의 난소와 자궁에서도 다량의 nesfatin-1/NUCB2가 발현된다는 사실이 새롭게 밝혀졌다. 그러나 생식기관에서 nesfatin-1/NUCB2 발현이 어떻게 조절되는지는 알려져 있지 않다. 따라서 본 연구에서는 생쥐의 생식기관 중 자궁에서 nesfatin-1/NUCB2 발현이 어떠한 경로를 통하여 조절되는지를 알아보고자 난소를

The Egr family of zinc finger transcription factors consisting of 4 members (Egr1 to Egr4) regulates critical genetic programs involved in cellular growth, differentiation, and function. They are co-ex-pressed in many different tissues, suggesting that they may have some redundant functions. While it is clear that estrogen regulates Egr1 in estrogen sensitive breast cancer cells, function of Egr1 and mechanisms by which estrogen (E2) and/or progesterone (P4) regulates Egr1 in uterus still remain unexplored. Thus, we have examined regulatory mechanisms by which Egr1 is regulated in the uterus and abnormal uterine phenotypes of Egr1(－/－) mice. Eight-week-old female mice were ovariectomized (OVX) and rested for a week. Uteri of OVX mice treated with various concentrations of E2 and/or other hormones were collected at 2 h after hormone treatment unless otherwise indicated. ICI 182,780 [estrogen receptor (ER) antagonist] or RU486 [progesterone receptor (PR) antagonist] was injected to OVX mice 30 min prior to hormone treatments. OVX Egr1(+/+) and Egr1(－/－) mice were treated with E2 and/or P4 to examine expression patterns of genes important for estrogen responses, and steroid hormone-induced cell proliferation in the uterus. Collected uteri were utilized for RT-PCR, realtime RT-PCR, Western blotting and histological analyses. Egr1 mRNA was rapidly induced with the highest level at 2h after E2 treatment and gradually deceased to basal levels at 12 h. Pretreatment of ICI 182,780 significantly reduced E2-induced increase of Egr1. However, an agonist for GPR30, a membrane estrogen receptor failed to induce mRNA expression of Egr1, suggesting that E2-dependent Egr1 transcription is mainly regulated via nuclear estrogen receptor, ER. P4 effectively dampened E2-dependent Egr1 transcription and its antagonistic effects were partially interfered with RU486 pretreatment. Histological analyses with BrdU incorporation experiments showed that vascular permeability (an early estrogen response) but not cell proliferation (a late response) was significantly impaired in the uteri of E2 treated OVX Egr1(－/－) mice. Interestingly, some genes involved in early estrogen responses such as Bip and HIF-1a but not those in late responses are dysregulated in uteri of Egr1(－/－) mice. Collectively, our results show that E2 transiently induces Egr1 via activation of nuclear ER. P4 antagonizes E2-dependent Egr1 regulation via PR. Impaired early estrogenic responses in Egr1(－/－) uteri could be due to aberrant gene expression affected by loss of Egr1 which act as a master regulator of estrogen actions in the uterus.-ex

The tremendous changes of uterine endometrium are observed during early pregnancy and protease and their inhibitors are involved in regulation of cell proliferation and remodeling of the tissues through remodeling the extracellular matrix (ECM). Some of the proteases and protease inhibitors have been suspected to a factor in endometrial changes but many parts of their expression profiles and the physiological roles are not uncovered. To evaluate the functional roles of them, in this study the expression profiles of proteases and protease inhibitors were analyzed using real-time quantitative PCR analysis. Mmp9 (matrix metalloproteinase 9) mRNA levels peaked on day 4 at the time of implantation. On the other hand, Ela2 (neutrophil elastase, NE) mRNA levels were peaked on day 2 of pregnancy. Its expression were decreased until day 4 of pregnancy but increased rapidly until day 7 of pregnancy and decreased again. NE inhibitor Slpi (secretory leukocyte protease inhibitor, SLPI) mRNA levels were related with the implantation stage and with the levels of Ela2. At the time of implantation the expression levels of Slpi mRNA were about 5 times higher than the Ela2 mRNA in the uterus. In the implantation stage embryos, Mmp9 specific mRNA was only detected at the blastocyst. On the other hand, the expression level of SLPI was higher than that of the Ela2 mRNA at blastocyst and 4.5 day p.c. embryos. Based on these results it is suggested that MMP9, SLPI, and NE have important physiological role in embryo implantation both in uterus and embryos.

착상기 이전 자궁에서 특이적 자궁내막 준비가 진행되어야 하는데, 이는 자궁 내막의 점진적 분화로 배아의 착상과 성공적 임신에 절대적으로 필요하다. 배아 발생 동안에 관찰되는 조직의 재구성은 세포외 기질을 포함한 다양한 요인에 의해 조절된다. 임신 동안에 관찰되는 극적인 변화로는 배아의 이동, 탈락막 반응, 태반의 분화를 그 예로 들 수 있다. 배아와 자궁간의 성공적 착상을 위한 변화들은 배아와 자궁의 착상을 위한 능력 갖출 수 있도록 한다. 이러한 변화

Aquaporins(AQPs)유전자는 다양한 조직의 상피세포와 내피세포에 존재하며 다량의 물 이동을 조절하는 막성 단백질로서, 세포간 또는 세포막 사이의 물 이동에 중요한 역할을 하는 것으로 알려져 있다. 발정기의 생쥐 자궁에서는 자궁내막세포의 증식과 함께 수화되는 특징을 보이며, 자궁내강으로 물이 이동되어 자궁내액의 점성이 낮아지는 현상이 나타난다. 따라서, 본 연구에서는 생쥐의 발정주기 동안 자궁에서의 형태학적인 변화와 관련하여 AQP유전자가 물 이